Agglutination, the reaction of bivalent antibodies with multivalent antigens to produce aggregates that can be detected and measured in various ways, has long been used to detect and quantify a wide variety of materials present in biological fluids. These materials, known generally as analytes, include drugs, drug metabolites, serum proteins, enzymes, and other materials of interest or clinical significance. With appropriate variations of the procedure either specific antigens or specific antibodies can be detected.
Increased sensitivity can be achieved by the use of particle reagents. A particle reagent typically consists of a latex particle containing an analyte, analyte derivative, or analyte analog on its surface. Although the analyte can be absorbed onto the surface of the latex particle, desorption during storage or use can produce variations in reagent properties that affect the sensitivity and reproducibility of the assay. Desorption problems can be overcome by covalently attaching an analyte, analyte derivative, or analyte analog to the surface of the particle. However, particle reagents in which the analyte, analyte derivative, or analyte analog is covalently attached to the surface of the particle can preaggregate during manufacture or not perform well in the intended assay. Preaggregation is an especially severe problem for particle reagents in which the covalently attached group is derived from a hydrophobic analyte.
To overcome the problem of preaggregation, functionalized polymer particles, in which the analyte, analyte derivative, or analyte analog is covalently attached to the surface of the particle by a hydrophilic linking agent, have been prepared. Hydrophilic diamines or polyamines have been used to attach groups that are either inherently amine-reactive or can be made amine-reactive by activation to epoxide-functionalized latex particles. Frey and Simons, U.S. Pat. No. 4,581,337, for example, disclose attachment of a theophylline derivative to epoxide-functionalized latex particles with hydrophilic diamine or polyamine linking agents.
The stability of the reagent is generally affected by several factors, including the amount of negative charge on the particle and the relative balance between hydrophilic and hydrophobic regions on the particle surface. Inclusion of excess hydrophilic diamine or polyamine during preparation of the functionalized polymer particle produces particles whose surface has been modified by reaction with the excess amine. Particle reagents containing these particles perform well in immunoassays, showing, for example, a rapid and precise agglutination rate when combined with an appropriate antibody.
In some cases, however, it is desirable to synthesize functionalized polymer particles from hydrophobic analytes that lack the amine-reactive functionality required for attachment by a hydrophilic diamine or polyamine linking agent. Particle reagents prepared with certain drugs or drug derivatives, particularly hydrophobic ones such as lidocaine, quinidine, procainamide, and derivatives of procainamide such as formyl procainamide, tend to aggregate during synthesis in the presence of excess hydrophilic diamine or polyamine, making them unusable in turbidimetric immunoassays. Thus, a need exists for stabilized polymer particles suitable for use in immunoassays that are not susceptible to premature aggregation.